Recent evidence suggests that the synthesis of 5-hydroxytryptamine (5HT) in the mammalian central nervous system like that of norepinephrine (NE) is enhanced by nerve stimulation, and depressed when intraneuronal levels of transmitter are raised by inhibiting monoamine oxidase. Not only are there these similarities between the regulatory mechanisms controlling the formation of the two monamines, but the rate limiting enzymes in the synthesis of NE and 5HT tyrosine hydroxylase and tryptophan hydroxylase also have certain properties in common, namely the requirement for a reduced pteridine cofactor and molecular oxygen. Regulation of tyrosine hydroxylase activity during short term nerve activity is effected in part by a reduction in feedback inhibition by NE. However, recent evidence now suggests that there may also be an allosteric activation of the enzyme by calcium. This is a highly significant finding because it is the entry of calcium into the nerve terminal during depolarization by nerve impulses that is believed to trigger the release process. The research proposal presented here is designed to test whether such an activation also occurs with tryptophan hydroxylase and if so, whether it would account for the increases in 5HT synthesis seen in intact tissues on nerve stimulation. In these experiments high speed supernatant preparations of tryptophan hydroxylase obtained from homogenates of rat brain will be assayed by measuring the formation of L-5-hydroxytryptophan from L-tryptophan using an assay in which the pteridine cofactor is regenerated by added pteridine reductase. The effects of calcium on the kinetic properties of the enzyme will be determined.